Jin Min, Park Seon-Joo, Kim Seok Won, Kim Hye Rim, Hyun Jin Won, Lee Jung-Hee
Laboratory of Genomic Instability and Cancer therapeutics, Cancer Mutation Research Center, Chosun University School of Medicine, Gwangju 61452, Republic of Korea.
Department of Cellular and Molecular Medicine, Chosun University School of Medicine, Gwangju 61452, Republic of Korea.
Biomol Ther (Seoul). 2017 Jul 1;25(4):396-403. doi: 10.4062/biomolther.2017.086.
Under normal, non-stressed conditions, intracellular p53 is continually ubiquitinated by MDM2 and targeted for degradation. However, in response to severe genotoxic stress, p53 protein levels are markedly increased and apoptotic cell death is triggered. Inhibiting the ubiquitination of p53 under conditions where DNA damage has occurred is therefore crucial for preventing the development of cancer, because if cells with severely damaged genomes are not removed from the population, uncontrolled growth can result. However, questions remain about the cellular mechanisms underlying the regulation of p53 stability. In this study, we show that p53-inducible gene 3 (PIG3), which is a transcriptional target of p53, regulates p53 stability. Overexpression of PIG3 stabilized both endogenous and transfected wild-type p53, whereas a knockdown of PIG3 lead to a reduction in both endogenous and UV-induced p53 levels in p53-proficient human cancer cells. Using both and ubiquitination assays, we found that PIG3 suppressed both ubiquitination- and MDM2-dependent proteasomal degradation of p53. Notably, we demonstrate that PIG3 interacts directly with MDM2 and promoted MDM2 ubiquitination. Moreover, elimination of endogenous PIG3 in p53-proficient HCT116 cells decreased p53 phosphorylation in response to UV irradiation. These results suggest an important role for PIG3 in regulating intracellular p53 levels through the inhibition of p53 ubiquitination.
在正常、无应激条件下，细胞内的p53会持续被MDM2泛素化并靶向降解。然而，在应对严重的基因毒性应激时，p53蛋白水平会显著升高并触发凋亡性细胞死亡。因此，在发生DNA损伤的情况下抑制p53的泛素化对于预防癌症发展至关重要，因为如果基因组严重受损的细胞没有从群体中被清除，就可能导致不受控制的生长。然而，关于p53稳定性调控的细胞机制仍存在问题。在本研究中，我们表明p53诱导基因3（PIG3）作为p53的转录靶点，可调节p53的稳定性。PIG3的过表达使内源性和转染的野生型p53均稳定，而敲低PIG3会导致p53功能正常的人类癌细胞中内源性和紫外线诱导的p53水平降低。通过使用体内和体外泛素化测定，我们发现PIG3抑制了p53的泛素化和MDM2依赖的蛋白酶体降解。值得注意的是，我们证明PIG3直接与MDM2相互作用并促进MDM2泛素化。此外，在p53功能正常的HCT116细胞中消除内源性PIG3会降低紫外线照射后p53的磷酸化。这些结果表明PIG3在通过抑制p53泛素化来调节细胞内p53水平方面发挥重要作用。
